Growth-limiting Conditions Research Articles

How carcinogens (or telomere dysfunction) induce genetic instability: associated-selection model

Carcinogens induce carcinogen-specific genetic instability (defects in DNA repair). According to the ‘direct-selection’ model, defects in DNA repair per se provide an immediate growth advantage. According to the ‘associated-selection’ model, carcinogens merely select for cells with adaptive mutations. Like any mutations, adaptive mutations occur predominantly in genetically unstable cells. The ‘associated-selection’ model predicts that carcinogen-driven selection minimizes cytotoxic but maximizes mutagenic effects of carcinogens. A purely mutagenic (neither cytotoxic, nor cytostatic) environment will favor effective DNA repair, whereas any growth-limiting conditions (telomerase deficiency, anticancer drugs) will select for genetically unstable cells. Genetic instability is a postmark of selective pressure rather than a hallmark of cancer per se. Once selected, genetic instability facilitates the development of resistance to any other growth-limiting conditions. As an example, a putative link between prior exposure to carcinogens and the ability to develop a telomerase-independent growth is discussed.

A Cell Population Model with Structured Medium for Budding Yeast Cultures

Continuous cultures of Saccharomyces ceremsiaeare known to exhibit sustained oscillations under aerobic and glucose limited growth conditions. The oscillations are reflected in measurements of both intracelluar and extracellular variables. Our recent work has shown that unstructured cell population balance models are capable of generating sustained oscillations over a range of dilution rates. A disadvantage of unstructured models is they lack variables that can be compared directly to experimental data. In this paper we present a structured description of the medium that allows the cell population model predictions to be compared to easily measured extracellular variables such as the dissolved oxygen and evolved carbondioxide concentrations. The proposed model is shown to replicate transient behavior observed experimentally for both batch and continuous cultures

Bifurcation Analysis for Discrimination of Budding Yeast Models

Continuous cultures of budding yeast are known to exhibit sustained oscillations under aerobic and glucose limited growth conditions. Many transient models have been proposed to explain this phenomenon. While they differ according to their underlying assumptions and mathematical complexity, all these models produce oscillations through bifurcations of the model equations. When combined with experimental data, bifurcation analysis provides a powerful tool to discriminate between alternative models. In this paper the bifurcation mechanisms of two representative yeast models are analyzed and compared with data. The two models investigated are: (i) an unsegregated cybernetic model in which oscillations arise due to competition between various metabolic pathways; and (ii) an unstructured cell population model in which oscillations arise due to cell cycle synchronization

The Autotrophic Synthesis of Polyhydroxyalkanoate by Alcaligenes eutrophusin the Presence of Carbon Monoxide

The CO-resistant strain B5786 of the hydrogen-oxidizing bacterium Alcaligenes eutrophuswas found to be able to synthesize polyhydroxyalkanoates (PHAs) under the conditions of growth limitation by nitrogen deficiency (the factor that promotes PHA synthesis) and growth inhibition by carbon monoxide. The gas mixtures that contained from 5 to 20 vol % CO did not inhibit the key enzymes of PHA synthesis–β-ketothiolase, acetoacetyl-CoA reductase, hydroxybutyrate dehydrogenase, and PHA synthase. In the presence of CO, cells accumulated up to 70–75 wt % PHA (with respect to the dry biomass) without any noticeable increase in the consumption of the gas substrate. Chromatographic–mass spectrometric analysis showed that the PHA synthesized by A. eutrophusis a copolymer containing more than 99 mol % β-hydroxybutyrate and trace amounts of β-hydroxyvalerate. The PHA synthesized under the conditions described did not differ from that synthesized by A. eutrophuscells from electrolytic hydrogen.

Epstein-Barr virus small RNAs potentiate tumorigenicity of Burkitt lymphoma cells independently of an effect on apoptosis.

The tumorigenic potential of the Burkitt lymphoma (BL) cell line Akata is dependent on the restricted latency program of Epstein-Barr virus (EBV) that is characteristically maintained in BL tumors. Within these cells, EBV-mediated inhibition of apoptosis correlates with an up-regulation of BCL-2 levels in concert with a down-regulation in c-MYC expression that occurs under growth-limiting conditions. Here we addressed whether EBV's effects on apoptosis and tumorigenicity are mediated by the EBV small RNAs EBER-1 and EBER-2. Stable expression of the EBERs in EBV-negative Akata BL cells, at levels comparable to those in EBV-positive cells, significantly enhanced the tumorigenic potential of EBV-negative BL cells in SCID mice, but did not fully restore tumorigenicity relative to EBV-positive Akata cells. Furthermore, wild-type or greater levels of EBER expression in EBV-negative Akata cells did not promote BL cell survival. These data therefore suggest that EBV can contribute to BL through at least two avenues: an EBER-dependent mechanism that enhances tumorigenic potential independent of a direct effect on apoptosis, and a second mechanism, mediated by an as-yet-unidentified EBV gene(s), that offsets the proapoptotic consequences of deregulated c-MYC in BL.

RpoS Mutants in Archival Cultures of Salmonella enterica Serovar Typhimurium

Long-term survival under limited growth conditions presents bacterial populations with unique environmental challenges. The existence of Salmonella enterica serovar Typhimurium cultures undisturbed in sealed nutrient agar stab vials for 34 to 45 years offered a unique opportunity to examine genetic variability under natural conditions. We have initiated a study of genetic changes in these archival cultures. We chose to start with examination of the rpoS gene since, among gram-negative bacteria, many genes needed for survival are regulated by RpoS, the stationary-phase sigma factor. In each of 27 vials examined, cells had the rpoS start codon UUG instead of the expected AUG of Salmonella and Escherichia coli strains recorded in GenBank. Ten of the 27 had additional mutations in the rpoS gene compared with the X77752 wild-type strain currently recorded in GenBank. The rpoS mutations in the 10 strains included two deletions as well as point mutations that altered amino acid sequences substantially. Since these stored strains were derived from ancestral cells inoculated decades ago and remained undisturbed, it is assumed that the 10 rpoS mutations occurred during storage. Since the remaining 17 sequences were wild type (other than in the start codon), it is obvious that rpoS remained relatively stable during decades of sealed storage.

Air pressure effects on biomass yield of two different Kluyveromyces strains.

The use of air pressure as a way of improving oxygen transfer in aerobic bioreactors was investigated. To compare the air pressure effects with traditional air bubbled cultures, experiments using a pressure reactor and a stirred flask, with the same oxygen transfer rate, were made. Kluyveromyces marxianus is an important industrial yeast and some of it show a "Kluyver effect" for lactose: even under oxygen limited growth conditions, certain disaccharides that support aerobic, respiratory growth, are not fermented. This study deals with the effect of increased pressure on the physiological behavior of two Kluyveromyces strains: K. marxianus ATCC10022 is a lactose-fermenting strain, whereas K. marxianus CBS 7894 has a Kluyver-effect for lactose. For K. marxianus ATCC10022 an air pressure increase of 2 bar led to a 3-fold increase in biomass yield. When air pressure increased an enhancement of ethanol oxidation of cell yeasts was also observed. Batch cultures of K. marxianus CBS 7894 exhibited different growth behaviour. Its metabolism was always oxidative and ethanol was never produced. With the increase in air pressure, it was possible to increase the productivity in biomass of K. marxianus CBS 7894. As a response to high oxygen concentrations, due to the increase in oxygen partial pressure, oxidative stress in the cells was also studied. Antioxidant defences, such as superoxide dismutase, catalase, and glutathione reductase, were at high activity levels, suggesting that these yeast strains could tolerate the increased pressures applied.

Vancomycin production in batch and continuous culture

Production of the glycopeptide antibiotic vancomycin by two Amycolatopsis orientalis strains was examined in batch shake flask culture in a semidefined medium with peptone as the nitrogen source. Different growth and production profiles were observed with the two strains; specific production (Y(p/x)) was threefold higher with strain ATCC 19795 than with strain NCIMB 12945. A defined medium with amino acids as the nitrogen source was developed by use of the Plackett-Burman statistical screening method. This technique identified certain amino acids (glycine, phenylalanine, tyrosine, and arginine) that gave significant increased specific production, whereas phosphate was identified as inhibitory for high specific vancomycin production. Experiments made with the improved medium and strain ATCC 19795 showed that vancomycin production kinetics were either growth dissociated or growth associated, depending on the amino acid concentration. In chemostat culture at a constant dilution rate (0.087 h(-1)), specific vancomycin production rate (q(vancomycin)) decreased linearly as the medium phosphate concentration was increased from 2 to 8 mM. In both phosphate and glucose limited chemostats, q(vancomycin) was a function of specific growth rate; the maximum value was observed at D = 0.087 h(-1) (52% of the maximum specific growth rate). Under phosphate limited growth conditions, q(vancomycin) was threefold higher (0.37 mg/g dry weight/h) than under glucose limitation (0.12 mg/g dry weight/h). (c) 1996 John Wiley & Sons, Inc.

Insulin-like signaling in yeast: modulation of protein phosphatase 2A, protein kinase A, cAMP-specific phosphodiesterase, and glycosyl-phosphatidylinositol-specific phospholipase C activities.

Previously, we have described significant effects of human insulin on glucose metabolism in the yeast Saccharomyces cerevisiae under conditions of growth limitation. These regulations apparently rely on a transmembrane receptor capable of binding human insulin and responding by tyrosine/serine phosphorylation of a specific set of polypeptides [Müller, G., Rouveyre, N., Crecelius, A., and Bandlow, W. (1998) Biochemistry 37, 8683-8695; Müller, G., Rouveyre, N., Upshon, C., Gross, E., and Bandlow, W. (1998) Biochemistry 37, 8696-8704; Müller, G., Rouveyre, N., Upshon, C., and Bandlow, W. (1998) Biochemistry 37, 8705-8713]. To characterize the molecular link between the initial steps in insulin-like signaling in yeast and the changes in the activities of glycogen synthase and glycogen phosphorylase, we examined here the effects of human insulin on a set of key regulatory enzymes of glycogen metabolism, protein phosphatase 2A (PP2A), cAMP-specific phosphodiesterase (cAMP-PDE), and protein kinase A (PKA). PP2A was activated about 2-fold by insulin in spheroplasts and in intact cells, whereas the fraction of active PKA was significantly reduced in a cAMP-independent manner as well as through a subsequent up to 3-fold increase in particulate cAMP-PDE activity accompanied by a 50% decrease in cytosolic cAMP levels. In addition, glycosyl-phosphatidylinositol-specific phospholipase C (GPI-PLC), which in isolated rat adipocytes is activated by insulin, was stimulated to up to 5-fold by glucose and 10-fold by glucose plus insulin in both yeast spheroplasts and intact cells leading to a concentration-dependent leftward shift of the glucose-response curve for activation of the GPI-PLC. GPI-PLC was most pronouncedly stimulated by authentic human insulin compared to various insulin analogues and insulin-like growth factor I. In addition to lipolytic cleavage by GPI-PLC, the GPI anchor of the cAMP-binding ectoprotein, Gce1p, was secondarily processed by a rapid proteolytic event. As the GPI-PLC reaction is rate limiting, the efficiency of the two-step anchor cleavage was significantly increased when insulin was present together with glucose as compared to glucose alone. The insulin concentrations effective in modulating PP2A, PKA, cAMP-PDE, and GPI-PLC activities correlate well with those required for half-saturation of the specific binding sites as well as for stimulation of protein phosphorylation and glycogen accumulation. The data suggest that mammalian insulin-sensitive cells and yeast share (part of) the key regulatory mechanism (consisting of PP2A, PKA, cAMP-PDE, and GPI-PLC) involved in the transduction of the insulin signal from the respective receptor systems to glycogen synthase and phosphorylase.

Carotenoid content of chlorophycean microalgae: factors determining lutein accumulation in Muriellopsis sp. (Chlorophyta)

Fifteen strains of chlorophycean microalgae have been investigated with regard to their carotenoid profile. Lutein, β-carotene and violaxanthin were present in virtually all of the strains, lutein, in general, being the most abundant carotenoid, whereas canthaxanthin and astaxanthin were found in some strains only. Chlorella fusca SAG 211-8b, Chlorococcum citriforme SAG 62.80, Muriellopsis sp., Neospongiococcum gelatinosum SAG B 64.80 and Chlorella zofingiensis CCAP 211/14 exhibited high lutein levels, the latter strain containing in addition substantial amounts of astaxanthin. Muriellopsis sp. was further characterized, since besides a high lutein content (up to 35 mg l −1 culture), it had the highest growth rate (up to 0.17–0.23 h −1) and maximal standing cell density (up to 8×10 10 cells l −1 culture). These levels of lutein are in the range of those reported for astaxanthin in Haematococcus and for β-carotene in Dunaliella, microalgae of recognized interest for the production of these carotenoids. Lutein content of Muriellopsis sp. increased during the exponential phase of growth, with the highest value being recorded in the early stationary phase. Maximum levels of lutein in Muriellopsis sp. cultures were recorded at 20–40 mM NaNO 3, 2–100 mM NaCl, 460 μmol photon m −2 s −1, pH 6.5 and 28°C, conditions which were, in general, also optimal for cell growth. Growth-limiting conditions, such as pH values of 6 or 9 and a temperature of 33°C, were found to stimulate carotenogenesis in Muriellopsis sp. This strain represents a potential source of lutein, a commercially interesting carotenoid of application in aquaculture and poultry farming, as well as in the prevention of cancer and diseases related to retinal degeneration.

Biotin production under limiting growth conditions by Agrobacterium/Rhizobium HK4 transformed with a modified Escherichia coli bio operon.

The E. coli biotin (bio) operon was modified to improve biotin production by host cells: (a) the divergently transcribed wild-type bio operon was re-organized into one transcriptional unit; (b) the wild-type bio promoter was replaced with a strong artificial (tac) promoter; (c) a potential stem loop structure between bioD and bioA was removed; and (d) the wild-type bioB ribosomal binding site (RBS) was replaced with an artificial RBS that resulted in improved bioB expression. The effects of the modifications on the bio operon were studied in E. coli by measuring biotin and dethiobiotin production, and bio gene expression with mini-cells and two-dimensional polyacrylamide gel electrophoresis. The modified E. coli bio operon was introduced into a broad host-range plasmid and used to transform Agrobacterium/Rhizobium HK4, which then produced 110 mg L-1 of biotin in a 2-L fermenter, growing on a defined medium with diaminononanoic acid as the starting material. Biotin production was not growth-phase dependent in this strain, and the rate of production remained high under limiting (maintenance) and zero growth conditions.

CO2 and light effects on deciduous trees: growth, foliar chemistry, and insect performance.

This study examined the effects of CO2 and light availability on sapling growth and foliar chemistry, and consequences for insect performance. Quaking aspen (Populus tremuloides Michx.), paper birch (Betula papyrifera Marsh.), and sugar maple (Acer saccharum Marsh.) were grown in controlled environment greenhouses under ambient or elevated CO2 (38.7 and 69.6 Pa), and low or high light availability (375 and 855 μmol m-2 s-1). Because CO2 and light are both required for carbon assimilation, the levels of these two resources are expected to have strong interactive effects on tree growth and secondary metabolism. Results from this study support that prediction, indicating that the relative effect of rising atmospheric CO2 concentrations on the growth and secondary metabolism of deciduous trees may be dependent on light environment. Trees in ambient CO2-low light environments had substantial levels of phytochemicals despite low growth rates; the concept of basal secondary metabolism is proposed to explain allocation to secondary metabolites under growth-limiting conditions. Differences between CO2 and light effects on the responses of growth and secondary metabolite levels suggest that relative allocation is not dependent solely on the amount of carbon assimilated. The relative growth rates and indices of feeding efficiency for gypsy moth (Lymantria dispar L.) larvae fed foliage from the experimental treatments showed no significant interactive effects of light and CO2, although some main effects and many host species interactions were significant. Gypsy moth performance was negatively correlated with CO2- and light-induced increases in the phenolic glycoside content of aspen foliage. Insects were not strongly affected, however, by treatment differences in the nutritional and secondary chemical components of birch and maple.

Threonine phosphorylation of modulator protein RsbR governs its ability to regulate a serine kinase in the environmental stress signaling pathway of Bacillus subtilis

The σ B transcription factor of the bacterium Bacillus subtilis controls the synthesis of over 100 general stress proteins that are induced by growth-limiting conditions. Genetic evidence suggests that RsbR modulates the phosphorylation state of the RsbS antagonist in the signaling pathway that regulates σ B activity in response to environmental stresses that limit growth. According to the current model, the phosphorylated RsbS antagonist is unable to complex RsbT, which is then released to initiate a signaling cascade that ultimately activates σ B. Here, we show that the RsbR protein itself has no kinase activity but instead stimulates RsbS phosphorylation by the RsbT serine kinase in vitro. We further show that in addition to its previously known serine kinase activity directed toward the RsbS antagonist, purified RsbT also possesses a threonine kinase activity directed toward residues 171 and 205 of the RsbR modulator. Threonine residues 171 and 205 were each found to be important for RsbR function in vivo, and phosphorylation of these residues abolished the ability of RsbR to stimulate RsbT kinase activity in vitro. These results are consistent with a model in which RsbR modulates the kinase activity of RsbT directed toward its RsbS antagonist in vivo, either specifically in response to environmental signals or as part of a feedback mechanism to prevent continued signaling.

Epstein-barr virus regulates c-MYC, apoptosis, and tumorigenicity in Burkitt lymphoma.

Loss of the Epstein-Barr virus (EBV) genome from Akata Burkitt lymphoma (BL) cells is coincident with a loss of malignant phenotype, despite the fact that Akata and other EBV-positive BL cells express a restricted set of EBV gene products (type I latency) that are not known to overtly affect cell growth. Here we demonstrate that reestablishment of type I latency in EBV-negative Akata cells restores tumorigenicity and that tumorigenic potential correlates with an increased resistance to apoptosis under growth-limiting conditions. The antiapoptotic effect of EBV was associated with a higher level of Bcl-2 expression and an EBV-dependent decrease in steady-state levels of c-MYC protein. Although the EBV EBNA-1 protein is expressed in all EBV-associated tumors and is reported to have oncogenic potential, enforced expression of EBNA-1 alone in EBV-negative Akata cells failed to restore tumorigenicity or EBV-dependent down-regulation of c-MYC. These data provide direct evidence that EBV contributes to the tumorigenic potential of Burkitt lymphoma and suggest a novel model whereby a restricted latency program of EBV promotes B-cell survival, and thus virus persistence within an immune host, by selectively targeting the expression of c-MYC.

Fabrication and physical properties of c-axis oriented thin films of layered perovskite La2−2xSr1+2xMn2O7

We have grown c-axis oriented films of a layered perovskite La2−2xSr1+2xMn2O7 (x=0.4) by pulsed laser deposition under the limited growth condition; above 900 °C and below 100 mTorr for substrate temperature and oxygen pressure (PO2), respectively. Otherwise, epitaxial but composition-unidentified films were deposited. The films show a resistive transition around 100 K in coincidence with the magnetic transition. The value of resistivity at low temperature is larger than that of a single crystal by about two orders of magnitude, perhaps due to the canted spin ordering. The films show gigantic magnetoresistance accompanied with hysteresis.

Strategies for the continuous production of ligninolytic enzymes in fixed and fluidised bed bioreactors

Production of ligninolytic enzymes by white-rot fungi is a phenomenon typically associated to the secondary metabolism, the main drawback of which is the limited yield of the enzyme obtained under growth-limiting conditions and the morphological behaviour of these filamentous microorganisms while growing. An efficient production system is needed for the promising applications of these enzymes in bioprocesses such as detoxification of effluents and biobleaching of kraft pulp. Several bioreactor configurations were considered which were operated in batch or semi-continuous modes. These operational conditions permitted high titres of enzymatic activity, however, the extent of production was limited. A different modus operandi is then to be proposed to overcome these limitations. The present work reviews the state of production of ligninolytic enzymes and proposes a new method to achieve continuous production of manganese peroxidase (MnP). The method comprises a feeding policy based on controlled feed rates of nutrients and the use of an adequate bioreactor configuration. They were both key factors to attain and sustain continuous production of MnP. The feeding of limited concentrations of nutrients kept secondary metabolism in steady-state conditions and limited the fungal growth. The configuration of two conventional bioreactors (fluidised bed and fixed bed bioreactors) was modified with the introduction of pulsation of gas. Pulsation proved to be a good alternative to enhance nutrients and oxygen transfer as well as to maintain fungal cultures as independent bioparticles both for pellets and foam blocks of mycelia. Other environmental factors such as a Mn 2+ concentration of 5 mM and high oxygen tension enhanced MnP production. The hydraulics of the bioreactor also had effect on MnP production, the optimum corresponded to a plug flow model with partial mixing and an operating HRT of 24 h. This policy facilitated long operation periods, obtaining higher productivity rates than the best reported to date.

CONSERVATION OF BANANA GERMPLASM THROUGH CRYOPRESERVATION

Since most banana landraces (Musa spp.) do not produce seed and are vegetatively propagated, germplasm must be maintained clonally. Therefore field and in vitro collections were established. Currently, the world's largest banana collection (1080 accessions) is kept as in vitro proliferating meristems under limited growth conditions at the laboratory of Tropical Crop Improvement of K.U.Leuven (Katholieke Universiteit Leuven, Belgium). Cryopreservation is a welcome alternative since during storage at ultra-low temperatures (-196°C), all metabolical, chemical and physical processes are arrested. As such, the material under storage does not need to be subcultured, thus avoiding risks of contamination and human error, and no somaclonal variation will take place. We developed an extremely simple cryopreservation system relying on a two-week-preculture phase of highly proliferating meristems on media with elevated sugar concentrations followed by rapid freezing in liquid nitrogen. This cryopreservation protocol was successfully tested on 20 accessions belonging to different genomic groups. Normal plants were regenerated from each of them. Regeneration frequencies vary from 7.4 to 68.9 %, depending on the cultivar. The mode of action of the sugar preculture phase with respect to the cryopreservation ability of different cultivars is under investigation. It is demonstrated that lipid, sugar and protein contents of non-precultured and sugar precultured plants differ.

Insulin signaling in the yeast Saccharomyces cerevisiae. 1. Stimulation of glucose metabolism and Snf1 kinase by human insulin.

Effects of human insulin on glucose metabolism in the yeast Saccharomyces cerevisiae were studied in this report. Under two conditions of growth limitation (glucose-grown cells during transition to stationary phase or spheroplasts during incubation in synthetic glucose medium), human insulin (10 and 1 microM, respectively) enhanced glycogen accumulation and glycogen synthase activity by 40-60% compared to control cells. Glycogen phosphorylase activity was also increased under the same conditions, but this stimulation was diminished by 35-45% in insulin-treated compared to control cells. Thus, under growth limitation, insulin causes glycogen phosphorylase and glycogen synthase to become more sensitive to inactivation and activation, respectively. In glucose-induced spheroplasts, insulin (1 microM), in addition to glycogen accumulation, led to about 2-fold increases of the rates of ethanol production and glucose oxidation compared to control cells, and the maximal concentration of hexose 6-phosphate was increased by 30-40%. In contrast, glucose transport as well as the levels of the allosteric regulators, fructose 2,6-bisphosphate and cAMP, were not altered at all. Snf1 kinase is assumed to be involved in the regulation of glycogen metabolism in yeast, although it does not seem to be modulated directly by the glucose concentration. Snf1 kinase activity was elevated 5-10-fold in response to insulin both during glucose induction of yeast spheroplasts and during transition to stationary phase of glucose-grown cells. We conclude that Saccharomyces cerevisiae and insulin-sensitive mammalian cells share some parts of the signaling cascades regulating oxidative and nonoxidative glucose metabolism in response to glucose and insulin.

The effect of Al on Ga desorption during gas source-molecular beam epitaxial growth of AlGaN

In this letter, we report on the impact aluminum has on gallium desorption kinetics in AlGaN alloys grown by gas source-molecular beam epitaxy. Aluminum is found to preferentially incorporate into the AlGaN films over the range of fluxes and temperatures investigated [0.05⩽Ji(Ga)⩽0.5 ML/s; 0.1⩽Ji(Al)⩽0.2 ML/s; 700 °C⩽Ts⩽775 °C]. As a result, Ga is not observed to incorporate into the film until the NH3 flux exceeds that required to grow stoichiometric AlN. This preferential incorporation stems from two facts: (a) Al has an ammonia cracking efficiency ∼2.5 times greater than that of Ga, and (b) Al participates in a Al-for-Ga exchange. As a result of these factors and under NH3 limited growth conditions, the aluminum mole fraction in a layer can be controlled by changing the incident NH3 flux.

The requirements for G1 checkpoint progression of Trypanosoma brucei S 427 clone 1

Trypanosoma brucei S 427 clone 1 accumulated in G1 when incubated under growth-limiting conditions. Further incubation of the G1-restricted organisms in medium containing 10% fetal bovine serum (FBS) and 2 mM hydroxyurea resulted in their reversible arrest after a G1 checkpoint beyond which serum was not required for progress into and through S. Progress of the G1-restricted T. brucei through the G1 checkpoint was linear and required continuous incubation with exogenous serum growth factors. These were principally low and high density lipoproteins; both lipoproteins triggered G1 progression in a dose- and time-dependent manner whilst their removal by immunoaffinity chromatography severely reduced the capacity of FBS to stimulate G1 progression. Serum-induced progress of T. brucei through G1 was Ca2+-independent, but required gene transcription, protein synthesis, and continuous kinase activity that was inhibited by tyrphostin 51 and DAPH 1 which typically inhibit epidermal growth factor receptor protein tyrosine kinase activity. The tyrphostin 51-sensitive catalytic activity was not required for T. brucei protein synthesis, glycolysis, or S phase progression but was required for tyrosine phosphorylation of several polypeptides, none of which was specifically associated with serum-induced G1 progression.